การผลิตเสนใยโปรตีนไหมระดับนาโนเมตรโดยเทคนิคการปนเสนใยดวยไฟฟาสถิต

Electrostatic Spinning of Silk Fibroin Nanofibers

ชิดชนก มีใจซือ่ ,1* รุจิรา วราพุฒ,1 ธนพล หวงจริง,1 นันทนา เขตบํารุง,1 และ พิชญ ศุภผล 2
Chidchanok Meechaisue,1* Rujira Waraput,1 Thanapol Huangjing,1 Nantana Ketbomrung,1
and Pitt Supaphol 2
1
Department of Materials Technology, Faculty of Science, Ramkhamhaeng University,
Bangkok 10240, Thailand
2
Technological Center for Electrospun Fibers, The Petroleum and Petrochemical College,
Chulalongkorn University, Bangkok 10330, Thailand

บทคัดยอ: กระบวนการปนเสนใยดวยไฟฟาสถิตเปนกระบวนการที่อาศัยแรงทางไฟฟาในการผลิตเสนใยที่
มีความละเอียดสูง ที่ซึ่งมีขนาดเสนผาศูนยกลางอยูในชวงตั้งแตระดับนาโนเมตรจนถึงระดับต่ํากวาสิบ
ไมโครเมตร เสนใยอิเล็กโทรสปนซึ่งจะเกิดขึ้นในลักษณะซอนทับกันเปนแผนเสนใยที่ไมไดทอมีลักษณะ
เดนที่นาสนใจหลายประการ เชน อัตราสวนของพื้นที่ผิวตอปริมาตรที่สูงมาก และขนาดของรูที่เกิดขึ้น
ระหวางเสนใยที่ซอนทับกันที่ เล็กมาก ทําใหเหมาะที่จะนําเสนใยอิเ ล็กโทรสปนเหลานี้ ไปใชงานดาน
การแพทยได ในการศึกษานี้ นํากระบวนการปนเสนใยดวยไฟฟาสถิตมาใชในการขึ้นรูปเสนใยนาโนจาก
โปรตีนไหมจากสารละลายโปรตีนไหมในกรดฟอรมิก ผลของความเขมขนของสารละลายที่มีตอสัณฐาน
วิทยาและขนาดของเสนใยที่ไดถูกศึกษาดวยกลองจุลทรรศนอิเล็กตรอนแบบสองกราด โดยทั่วไป เสนใย
โปรตีนไหมที่ผลิตไดมีขนาดใหญขึ้นเมื่อความเขมขนของสารละลายมีคาเพิ่มมากขึ้น

Abstract: Electrostatic spinning or electrospinning is a process in which the electrical forces

are responsible for the production of ultra-fine fibers with diameters in the range of
nanometers to sub-micrometers. The electrospun fibers formed as non-woven mats exhibit
several interesting characteristics, for examples, large surface area-to-volume ratio, small
pore size between depositing fibers of the fiber mats, making these electrospun fibers suitable
for use in biomedical applications. In this study, electrospinning was used to prepare silk
fibroin nanofibers from solutions of silk fibroin in formic acid. The effect of solution
concentration on morphological appearance and size of the obtained fibers was investigated
by electron scanning microscopy (SEM). Generally, the fiber diameters were found to
increase with increasing concentration of the silk concentration.

Introduction: Electrostatic spinning or electrospinning is a process by which ultrafine fibers

with diameters in the sub-micrometer down to nanometer range can be produced. The
manufacture of these ultrafine fibers concerns with the application of a high electrostatic
potential from an emitting electrode of a high-voltage power supply to the polymer solution
or melt across a finite distance between a conductive nozzle and a grounded collective
screen. The ultrafine fibers from this electrospinning process exhibit several interesting
characteristics, for examples, large surface area to mass or volume ratio, small pore size
between depositing fibers of the electrospun mat, and flexibility for surface functionalization.

31st Congress on Science and Technology of Thailand at Suranaree University of Technology, 18 – 20 October 2005
These unique properties render electrospun ultrafine fibers as excellent candidates for a
number of biomedical applications: for examples, tissue scaffolding materials, wound healing
materials, vascular grafts, and drug delivery carriers.
Besides using natural silk as high-quality textile fibers, many researchers have recently
investigated silk as one of candidate materials for biomedical applications because it has
several useful properties including good biocompatibility, biodegradability, good oxygen and
water vapor permeability, and minimal inflammatory reaction. The natural silk fibers
Bombyx mori consist of two types of proteins, fibroin and sericin. Fibroin is the protein that
forms the filaments of silk-worm silk, whereas sericin is, in fact, a group of gummy proteins
that bind the fibroin filaments. Silk fibroin can be used in various forms, such as gels,
powders, fibers, or membranes, depending on applications.
In this work, silk fibroin (SF) was used to produce ultrafine fibers by the electrospinning
process. The effects of solution concentration and electrostatic field strength on
morphological appearance and size of the electrospun SF fibers were investigated using
scanning electron microscopy (SEM).

Methodology: Cocoons of Bombyx mori silkworm silk were boiled in water and then dried at
60oC for 24 hr in an oven to obtain raw silk fibers. These fibers were degummed three times
with 0.5% w/v Na2CO3 solution at 100oC for 30 min and then rinsed with warm water.
Degummed silk (silk fibroin, SF) was dissolved in a ternary solvent system of
CaCl2/CH3CH2OH/H2O (1:2:8 in mol ratio) at 70oC. This solution was dialyzed in water
using cellulose tubular membrane (Sigma-Aldrich, USA) for 3 days. SF solution was filtered
and lyophilized to obtain SF sponges. SF solutions for electrospinning were prepared by
dissolving weighed amount of SF sponges in 85% formic acid (Carlo Erbra, Italy) in various
concentrations (i.e. 10, 15, 20, 25, 30, 35 and 40% w/v). To electrospin SF fibers, each
solution was placed in a 5-ml glass syringe connected with a gauge 26 stainless steel needle.
A piece of aluminum sheet was used as the collective screen. A Gamma High Voltage
Research D-ES30PN/M692 power supply (Florida, USA) was used to generate high
electrostatic potentials. The applied electrostatic potential and the collection distance were
fixed at 20 kV and 10 cm, respectively. The morphological appearance of the as-spun SF
fibers was observed by a JEOL JSM-6400 scanning electron microscope (SEM).

Results, Discussion and Conclusion: Figure 1 shows selected SEM images of as-spun SF
fibers from SF solutions in various concentrations (i.e. 10, 15, 20, 25, 30, 35 and 40% w/v).
At low concentrations (e.g. 10% w/v), a large number of droplets were present (see Figure
1a). At such low concentrations, the viscoelastic force (i.e. a result of the low degree of
chain entanglements) was comparatively smaller than the Coulombic force. This resulted in
the over-stretching of a charged jet, hence the break-up of the charged jet into many small
droplets as a result of the surface tension. On the contrary, at higher concentrations, the
viscoelastic force became larger in comparison with Coulombic force (due mainly to the
increased chain entanglements). The increase in the viscoelastic force was sufficient to
prevent a charged jet from breaking up into small droplets and to allow the electrostatic stress
to further elongate the jet which finally thins down the diameter of the jet. At 15 and 20%
w/v, a mixture between beaded fibers and smooth fibers was obtained (see Figure 1b). When
the concentration of the solution increased to 30% w/v, the beads disappeared altogether,
leaving only smooth ultrafine fibers, with the average diameter about 213 nm, on the target

31st Congress on Science and Technology of Thailand at Suranaree University of Technology, 18 – 20 October 2005
(see Figure 1c). With further increasing concentration (i.e. 35 and 40% w/v), the initial
amount of the solvents in a small segment of a charged jet decreased, rendering the charged
jet to “dry” much easier. In addition, the increased concentration enabled the charged jet to
withstand larger stretching force (from the Coulombic repulsion), resulting in the observed
larger diameters of the charged jet (ultimately, the as-spun fibers). The average fiber
diameter was found to increase from ca. 213 nm at 30% w/v to ca. 526 nm at 40% w/v.

(a) (b)

(c) (d)
Figure 1. Scanning electron micrographs of electrospun silk fibroin fibers prepared from
a) 10, b) 20, c) 30 and d) 40% w/v silk fibroin solutions (magnification = 3000×).

References:
(1) Gregory H. Altman, Frank Diaz, Caroline Jakuba, Tara Calabro, Rebecca L. Horan,
Jingsong Chen, Helen Lu, John Richmond, David L. Kaplan (2003) Biomaterials, 24,
401-416.
(2) So Hyun Kim, Young Sik Nam, Taek Seung Lee, and Won Ho Park (2003) Polymer
Journal, 35, 185-190.
(3) Chidchanok Mit-uppatham, Manit Nithitanakul, and Pitt Supaphol (2004)
Macromolecular Chemistry and Physics, 205, 2327-2338.

Keywords: Electrospinning; Nanofibers; Silk fibroin

31st Congress on Science and Technology of Thailand at Suranaree University of Technology, 18 – 20 October 2005